The effects of host factors ADP-ribosylation factor 4 (ARF4) and ADP-ribosylation factor 5 (ARF5) upon Zika virus (ZIKV) infection in vivo were characterized by construction of gene knockout mice via CRISPR-Cas9. Firstly, ARF5 and ARF4 genes were modified by the CRISPR-Cas9 system and then microinjected into the fertilized eggs of C57BL/6JGpt mice. Fertilized eggs were transplanted to obtain ARF4 or ARF5 knockout (ARF4KO or ARF5KO) mice, and ARF4/5 double knockout mice were achieved by the mating between ARF4KO and ARF5KO mice (ARF4KO/ARF5KO). Then, the mouse genotypes were identified by PCR to identify the positive knockout mice, and RT-qPCR was employed to examine the knockout efficiency. The mice were then infected with ZIKV and the blood and tissue samples were collected after 2, 4, and 6 days. RT-qPCR was then employed to determine the virus load, and hematoxylin-eosin staining was employed to observe the pathological changes in the tissue. The results showed that expected PCR bands were detected from ARF4KO^-/+, ARF5KO^-/-, and ARF4KO^-/+/ARF5KO^-/- mice, respectively. The results of mRNA transcription measurement indicated the significant knockdown of ARF4 by 37.8%-50.0% but not ARF5 in ARF4KO^-/+ compared with the wild-type mice. Meanwhile, complete knockout of ARF5 and no changes in ARF4 were observed in ARF5KO^-/- mice. Additionally, completed knockout of ARF5 and down-regulated mRNA level of ARF4 in the lung, kidney, and testis were detected in ARF4KO^-/+/ARF5KO^-/-mice in comparison with the wild-type mice. The virus load in the serum decreased in ARF4KO^-/+ mice, while it showed no significant change in ARF5KO^-/- or ARF4KO^-/+/ARF5KO^-/- mice compared with that in the wild type. Meanwhile, ARF4KO^-/+ mice showcased no significant difference in virus load in various tissues but attenuated pathological changes in the brain and testis compared with the wild-type mice. We successfully constructed ARF4KO and ARF5KO mice by CRISPR-Cas9 in this study. ARF4 rather than ARF5 is essential for ZIKV infection in vivo. This study provided animal models for studying the roles of ARF4 and ARF5 in ZIKV infection and developing antivirals.
Animals ; ADP-Ribosylation Factors/metabolism* ; Zika Virus Infection/genetics* ; Mice ; Mice, Knockout ; Zika Virus/genetics* ; Mice, Inbred C57BL ; CRISPR-Cas Systems ; Viral Load ; Male ; Female

Since Zika virus (ZIKV) has firstly been isolated in 1947, Uganda, outbreaks of Zika fever have been reported in many areas such as in Africa, Southeast Asia and America. Imported cases in China also have been reported. Zika virus belongs to the family Flaviviridae, genus Flavivirus, and include Africa subtype and Asia subtype. It is a mosquito-borne virus primarily transmitted by Aedes aegypti mosquitoes. Sexual transmission, Blood transmission and mother-to-fetus transmission were also reported. Zika virus can go though blood-brain barrier and infect central nervous system. Symptoms are generally mild and self-limited, but recent evidence suggests a possible association between maternal Zika virus infection and adverse fetal outcomes, such as congenital microcephaly, as well as a possible association with Guillain-Barré syndrome. Laboratorial Diagnosis includes nucleic acid detection, Serological test, and isolation of virus. Currently, no vaccine or medication exists to prevent or treat Zika virus infection. Preventive measures against Zika virus infection should be taken through prevention of mosquito bites and surveillance in epidemic area.
Aedes ; physiology ; virology ; Animals ; Humans ; Insect Vectors ; physiology ; virology ; Zika Virus ; genetics ; physiology ; Zika Virus Infection ; transmission ; virology

OBJECTIVETo clarify the possible association between the Zika virus (ZIKV) and microcephaly and understand where we are in terms of research and the debate on the causation between mild maternal clinical features and severe fetal microcephaly.

DATA SOURCESWe did a comprehensive literature review with the keywords "zika" and/or "microcephaly" from inception to May 27, 2016, with PubMed.

STUDY SELECTIONStudies were included and analyzed if they met all of the following criteria: "probable or confirmed infant microcephaly" and "probable or confirmed ZIKV infection among mothers or infants".

RESULTSWe emphasize the diagnosis of ZIKV infection, including maternal clinical manifestations, maternal and fetal laboratory confirmation, and possible autopsy if need. Other confounders that may lead to microcephaly should be excluded from the study. We presented the results from clinical manifestations of ZIKV infection, testing methods evolving but the mechanism of microcephaly uncertain, flexible definition challenging the diagnosis of microcephaly, and limited causal reference on pregnant women. We made analog comparison of severe acute respiratory syndrome and chikungunya virus in terms of DNA mutation and global movement to provide further research recommendation. The chance of catch-up growth may decrease the number of pervious "diagnosed" microcephaly.

CONCLUSIONSThere are some evidence available through mice models and direct isolation of ZIKV in affected pregnancies on kindly causal relationship but not convincible enough. We analyzed and presented the weakness or limitation of published reports with the desire to shed light to further study directions.

Animals ; Female ; Humans ; Microcephaly ; diagnosis ; etiology ; genetics ; Mutation ; Pregnancy ; Zika Virus ; pathogenicity ; Zika Virus Infection ; complications

Since Zika virus has been spreading rapidly in the Americas from 2015, the outbreak of Zika virus infection becomes a global health emergency because it can cause neurological complications and adverse fetal outcome including microcephaly. Here, we report clinical manifestations and virus isolation findings from a case of Zika virus infection imported from Brazil. The patient, 43-year-old Korean man, developed fever, myalgia, eyeball pain, and maculopapular rash, but not neurological manifestations. Zika virus was isolated from his semen, and reverse-transcriptase PCR was positive for the virus in the blood, urine, and saliva on the 7th day of the illness but was negative on the 21st day. He recovered spontaneously without any neurological complications. He is the first case of Zika virus infection in Korea imported from Brazil.
Adult ; Brazil ; Humans ; Male ; Microscopy, Electron, Transmission ; RNA, Viral/analysis/blood/urine ; Republic of Korea ; Reverse Transcriptase Polymerase Chain Reaction ; Saliva/virology ; Semen/virology ; Travel ; Zika Virus/genetics/*isolation & purification ; Zika Virus Infection/*diagnosis/virology

Aedes ; virology ; Animals ; China ; epidemiology ; Genetic Variation ; Genome, Viral ; Humans ; Multigene Family ; Viral Proteins ; genetics ; Zika Virus ; classification ; genetics ; isolation & purification ; Zika Virus Infection ; epidemiology ; virology